Long‐term follow‐up of tandem <scp>CD19</scp>/<scp>CD22 CAR T‐Cells</scp> in r/r <scp>B‐ALL</scp> patients with high‐risk features
Authors: Wei Cui, Xin‐Yue Zhang, Zheng Li, Hai‐Ping Dai, Jia Yin, Qing‐Ya Cui, Si‐Ning Liu, Li‐Qing Kang, Lei Yu, De‐Pei Wu, Xiao‐Wen Tang
Journal: American Journal of Hematology (2023)

CD19 chimeric antigen receptor (CAR) T-cells therapy has shown remarkable therapeutic effect in treating relapsed and refractory B acute lymphoblastic leukemia (r/r B-ALL). However, approximately 50% of patients experience relapse within a year due to limited CAR T-cell persistence or CD19 antigen loss.1, 2 Dual-targeting approaches have demonstrated impressive antitumor activity.3 And it has been proven to prevent CD19 negative relapse in preclinical studies.4 Although the previous single reports have showed the safety and improved response of tandem dual-targeting CAR T-cells, overall follow-up data are limited.5, 6 Therefore, we conducted a prospective clinical trial in a large cohort to investigate the safety and efficacy of tandem CD19/CD22 CAR T-cells strategy in r/r B-ALL patients. This trial was registered at ClinicalTrial.gov as no. NCT 03614858. The CD19/CD22 CAR is a tandem CAR molecular, consisting of a CD28 and OX40 costimulatory domain (Figure 1B). More details of the protocol are found in Appendix S1. From August 2018 to June 2021, a total of 47 patients were treated with CD19/CD22 CAR T-cells (Figure 1A, Table S1). The cohort's median age was 28 years old (range 6–56). All patients had failed multiple therapies, including chemotherapy, tyrosine kinase inhibitor (TKI) therapy, or adoptive cell therapy. One patient had previously received CD19 CAR T-cells therapy, and nine patients had relapsed after allogeneic stem cell transplantation (allo-HSCT). The median of bone marrow (BM) blasts prior to the lymphodepletion regimen were 32.5% (range, 0%–94.5%), and 27 patients (57.4%) had a high disease burden (20%–94.5%). Seven patients had extramedullary disease. Most patients had high-risk cytogenetic and/or molecular abnormalities, including BCR/ABL fusion (n = 18), Philadelphia chromosome-positive (Ph+) ALL with T315I mutation (n = 9), Ph-like ALL (n = 6), and TP53 alteration (n = 6). In addition, mutations such as PTPN11, ETV6, and SETD2 were observed by next-generation sequencing. All patients received an FC lymphodepletion regimen (fludarabine, 30 mg/m2 and cyclophosphamide, 300 mg/m2, on day −5 to −3). Then, patients received tandem CD19/CD22 CAR T-cells infusion according to the dose escalation schedule in 3 consecutive days (10% on day 1, 30% on day 2 and 60% on day3) (Figure 1C). The median infusion dose of CAR T-cells was 1 (0.5–2.5) × 107/kg. Details of adverse effects (AEs) are shown in Table S2. The most common treatment-related AE was cytokine release syndrome (CRS). It was mainly manifested as fever, hypotension, and respiratory distress. CRS of any grades occurred in 41 of 47 patients (87.2%) and were severe (grade >2) in 8 (17.0%). The above symptoms disappeared soon through the vasopressors, ventilatory support, and comprehensive support care in time. Immune effector cell-associated neurotoxicity syndrome was seen in one patient who had a high tumor burden (94.5% in BM). The patient also had grade 3 CRS, and the symptoms were relieved by low-dose dexamethasone (5 mg/day for 6 consecutive days), plasma exchange, and continuous renal replacement therapy. Hematologic toxic effects were the most common events of grade 3 or higher, including leukopenia (74.5%), anemia (48.9%), and thrombocytopenia (57.4%). Nonhematologic toxicities comprised infection (23.4%), liver injury (23.4%), and acute kidney injury (8.51%). Generally, all cases were well-tolerated and reversible. The primary response assessment was performed on day 28 after CAR T-cells infusion. The overall response rate was 100%. All 47 patients (100%) achieved hematological complete remission (CR). Forty patients (85.1%) got minimal residual disease negative (MRD−) CR by flow cytometry. Among these patients, 17 of 18 patients with Ph+ ALL, 6 patients with TP53 mutation, 7 patients with extramedullary involvement, 6 patients who relapsed after HSCT, and 3 of 6 patient with Ph-like ALL achieved MRD− CR. During a median follow-up of 29.6 months (range, 2.6–53.7), the median overall survival (OS) and leukemia-free survival (LFS) for the cohort have not reached. The OS rate was 76.2% (95% CI, 61.1%–86.1%) at 1 year and 73.8% (95% CI, 58.5%–84.2%) at 2 years in all patients (Figure 1D). A total of 12 patients relapsed after CAR T-cells therapy, with 10 having CD19+CD22+ relapse and 2 having CD19−CD22+ relapse, and the duration of remission was 9.7 months in these patients. The 2-year LFS rate and 2-year cumulative incidence of relapse (CIR) rate were 69.2% (95% CI, 53.6%–80.5%) and 22.7% (95% CI, 6.2%–45.2%), respectively (Figure 1E,F). After CAR T-cells infusion, 35 patients (74.5%) underwent a consolidative allo-HSCT. The median time to allo-HSCT was 2 months (range 1–5.5) from CAR T-cells infusion. After allo-HSCT, six patients relapsed and three died of disease progression. In addition, five patients died of transplantation-associated diseases. In no HSCT group, 8 of 12 patients had no bridging HSCT because they had HSCT history before CAR T-cells therapy. Four patients refused HSCT for personal reasons. Six out of 12 patients without transplantation relapsed and 3 died of disease progression. The HSCT group had significantly better LFS and lower CIR than the no HSCT group (LFS, p = .04; CIR, p = .02, Figure 1G–I). We initially performed univariable Cox regression analyses to identify the baseline and therapy-related elements that associated with better LFS. Univariable analysis and multivariate showed patients could benefit from bridging allo-HSCT. The baseline characteristics, such as age, high tumor burden before CAR-T, adverse genetics, and the previous therapies, were not strongly related to the duration of LFS (Table S3, Figure S2). In recent studies, dual-target CAR T-cells therapy has the potential to target an alternative antigen, besides CD19, so that it can mediate similar potent antineoplastic effects and triumph over antigen loss.4 Our study outcomes are very promising in high initial remission for high-risk patients. However, the efficacy of overcoming relapse is still not clear in the tandem CAR T-cells. The limitation of CAR T-cells persistence may account for the antigen positive relapse (Figure S3). As such, we herein for the first time summarized the clinical trials of CD19 CAR T-cells therapy in a large cohort and the CD19− relapse rate (5.3%–24.6%) in Table S4. A previous multicenter, retrospective study indicated that CD19− relapse accounted for 16.19% of 420 CAR-treated patients.7 In our study, two of 47 patients (4.3%) had CD19−CD22+ recurrence after CAR T-cells infusion and bridging transplantation. Unfortunately, we have not detected the CAR T-cells persistence during the relapse. The antigen negative relapse may be concerned regarding the functionality of the CD22 portion of CAR T-cells in vivo. Moreover, transplantation eliminating autologous origin CAR T-cells may account for the relapse. More fundamental studies are needed to explore this mechanism. Bridging allo-HSCT after tandem CD19/CD22 CAR T-cells therapy is an efficient and relatively safe way that prolongs LFS and reduces the relapse risk. An increasing number of clinical trials have compared the outcomes of patients with and without consolidative allo-HSCT after CD19 CAR-T therapy. A recent meta-analysis identified 19 clinical trials with 690 patients, and found that bridging HSCT was beneficial for OS, the relapse rate, and LFS.8 However, there is also a limitation in our study that the no HSCT group size was insufficient. Additional studies with larger sample numbers are needed to explore the possibility of transplantation free. In summary, tandem dual-targeting CD19/CD22 CAR T-cells therapy demonstrated a high CR rate in r/r B-ALL patients, regardless of their high-risk features, such as TP53 mutation, BCR-ABL fusion, or Ph-like ALL, as well as relapse after allo-HSCT. Consolidative allo-HSCT after the tandem CAR product could improve the long-term disease control in our study. W Cui and XY Zhang wrote the manuscript. LQ Kang and L Yu designed and prepared the CAR T-cells. XW Tang, HP Dai, J Yin, Z Li, and QY Cui treated the patients. XY Zhang and SN Liu collected and analyzed the primary data. DP Wu and XW Tang revised the manuscript. All authors approved the final version of the manuscript. This work was supported by research grants from National Natural Science Foundation of China (81873443, 82070162), Translational Research Grant of NCRCH (2020ZKZC04) and Natural Science Foundation of Jiangsu Province (BK20201169), The Key Science Research Project of Jiangsu Commission of Health (K2019022), Frontier Clinical Technical Project of Suzhou Science and Technology plan (SKY2022001), Bethune Charitable Foundation (BCF-IBW-XY-20220930-13), Suzhou diagnosis and treatment project of Clinical Key Diseases (LCZX202201), and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD). The authors declare that they have no potential conflict of interests. The data that support the findings of this study are available in the Appendix S1 of this article. Appendix S1. Supporting Information. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.